Comparative analysis of MACROD1, MACROD2 and TARG1 expression, localisation and interactome
Comparative analysis of MACROD1, MACROD2 and TARG1 expression, localisation and interactome
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DOI：
10.1038/s41598-020-64623-y
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发表时间：
2020-05-19
影响因子：
4.6
通讯作者：
Feijs, K. L. H.
中科院分区：
文献类型：
--
作者：
Zaja, R.;Aydin, G.;Feijs, K. L. H.
翻译后修饰ADP-核糖基化参与许多细胞过程，多聚和单（ADP-核糖基）化（分别为PAR-和MAR化）具有不同的作用。MACROD 1、MACROD 2和TARG 1的发现证明了细胞内MARylation的可逆性，MACROD 1、MACROD 2和TARG 1是三种能够逆转蛋白质和RNA MARylation的含有大结构域的酶。虽然这三种酶在体外具有相同的活性，但它们在细胞中的作用尚不清楚，并且发表的数据部分相互矛盾，可能是由于缺乏经过验证的试剂。我们开发了单克隆抗体来研究这些蛋白质，并分析了它们的组织分布和细胞内定位。MACROD 1在骨骼肌的线粒体中最普遍，MACROD 2定位于细胞核和细胞质，迄今为止仅在神经母细胞瘤细胞中发现，而更普遍表达的TARG 1存在于核质、核仁和应激颗粒中。MACROD 1的缺失或TARG 1的缺失分别导致线粒体或核仁形态的破坏，暗示它们对这些细胞器的重要性。为了开始阐明潜在的机制，我们使用BioID绘制了它们的相互作用组。相互作用物的细胞定位分别支持MACROD 1和TARG 1的线粒体、核仁和应激颗粒定位。基因本体分析表明MACROD 1和TARG 1参与了它们各自隔室中的RNA代谢。本文对水解酶的表达、定位和相互作用组的详细描述为今后更详细地研究其生理功能提供了坚实的基础。
The posttranslational modification ADP-ribosylation is involved in many cellular processes, with distinct roles for poly- and mono(ADP-ribosyl)ation (PAR- and MARylation, respectively). Reversibility of intracellular MARylation was demonstrated with the discovery of MACROD1, MACROD2 and TARG1, three macrodomain-containing enzymes capable of reversing MARylation of proteins and RNA. While the three enzymes have identical activities in vitro, their roles in cells are unclear and published data are partially contradictory, possibly due to a lack of validated reagents. We developed monoclonal antibodies to study these proteins and analysed their tissue distribution and intracellular localisation. MACROD1 is most prevalent in mitochondria of skeletal muscle, MACROD2 localises to nucleo- and cytoplasm and is found so far only in neuroblastoma cells, whereas the more ubiquitously expressed TARG1 is present in nucleoplasm, nucleolus and stress granules. Loss of MACROD1 or loss of TARG1 leads to disruption of mitochondrial or nucleolar morphology, respectively, hinting at their importance for these organelles. To start elucidating the underlying mechanisms, we have mapped their interactomes using BioID. The cellular localisation of interactors supports the mitochondrial, nucleolar and stress granule localisation of MACROD1 and TARG1, respectively. Gene ontology analysis suggests an involvement of MACROD1 and TARG1 in RNA metabolism in their respective compartments. The detailed description of the hydrolases' expression, localisation and interactome presented here provides a solid basis for future work addressing their physiological function in more detail.